Synthesis of 3H, 13C,2H3,15N and 14C-labelled SCH 466036, a histamine 3 receptor antagonist

Article abstract of DOI:10.1002/jlcr.3261

The synthesis of [₃H]SCH 466036, [Me-₃H₃]SCH 466036, [₁₃C,₂H₃,₁₅N]SCH 466036 and [₁₄C]SCH 466036 is described. [₃H]SCH 466036 was prepared in two steps via Raney Ni-catalysed exchange with tritiated water. [Me-₃H₃]SCH 466036 was prepared in a single step from [₃H]methyl iodide in 45% yield. [₁₃C,₂H₃,₁₅N]SCH 466036 was prepared in two steps from [₁₅N] hydroxylamine and [₁₃C,₂H₃]methyl iodide with an overall yield of 16%. [₁₄C]SCH 466036 was prepared in seven steps from [₁₄C]potassium cyanide in an overall yield of 13%.

Full text of DOI:10.1002/jlcr.3261

Research Article
Received 20 October 2014,
Revised 03 December 2014,
Accepted 14 December 2014
Published online in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/jlcr.3261
3
13 2
15
14
Synthesis of H, C, H , N and C-labelled
3
SCH 466036, a histamine 3 receptor
antagonist
D. Hesk,* S. Borges, R. Dumpit, S. Hendershot, D. Koharski, C. Lavey,
P. McNamara, and K. Voronin
3
3
13
2
15
14
3
The synthesis of [ H]SCH 466036, [Me- H ]SCH 466036, [ C, H , N]SCH 466036 and [ C]SCH 466036 is described. [ H]SCH
3
3
3
4
66036 was prepared in two steps via Raney Ni-catalysed exchange with tritiated water. [Me- H ]SCH 466036 was prepared
in a single step from [ H]methyl iodide in 45% yield. [ C, H , N]SCH 466036 was prepared in two steps from [ N]
hydroxylamine and [ C, H
3
3 13 2 15 15
3
1
3
2
14
3
]methyl iodide with an overall yield of 16%. [ C]SCH 466036 was prepared in seven steps from
1
4
[
C]potassium cyanide in an overall yield of 13%.
Keywords: tritium; deuterium; carbon-14; nitrogen-15; SCH 466036; H3 antagonist
Introduction
a batch of 5.5mCi at 99% radiochemical purity at a specific activity
of 324mCi/mmole was obtained. H NMR analysis (Figure 2)
3
SCH 466036, 1, is a potent and selective histamine 3 (H3)
receptor antagonist.
showed that 51% of the tritium was present in the expected 6-
position in the 2-aminopyridine ring with an additional 43%
present in the piperidine ring, α to the oxime, which is likely
1
–4
6,7
incorporated by a base-catalysed mechanism. The remaining
% was located in the monosubstituted ring adjacent to the
6
pyridine nitrogen, clearly showing a much lower affinity for the
Raney Ni catalyst than the 2-aminopyridine ring system.
3
High specific activity [ H]SCH 466036, for receptor binding
studies, was prepared by methylation of the free oxime 5 with
3
H3 antagonists have been shown to be effective in animal
models of allergy, congestion, obesity and metabolic
[
H]methyl iodide under phase-transfer conditions with the
phase-transfer catalyst ‘Aliquat 336’ (Figure 3).
1
–4
3
syndrome. [ H]SCH 466036 of moderate specific activity was
rapidly prepared to support preliminary absorption, distribution,
metabolism and excretion studies. In addition, high specific
A 10-fold excess of the free oxime 5 was used in an attempt to
3
optimize the utilization of the [ H]methyl iodide and thus
achieve a reasonable conversion to compound 6. The addition
3
activity [ H]SCH 466036 was prepared for H3 receptor-binding
of methylene chloride as an organic cosolvent facilitated the
3
studies. Subsequently, stable isotope-labelled SCH 466036 was
made for use as an internal standard to support the toxicology
solubility of the ketone in toluene, deriving from the [ H]methyl
iodide solution in toluene. After purification by reverse-phase
14
3
and clinical programmes, and [ C]SCH 466036 was then
prepared for the drug disposition group to conduct definitive
pharmacokinetic and drug metabolism studies. This paper
discusses the synthesis of all four isotopically labelled forms of
SCH 466036.
HPLC, a 45 mCi batch of [Me- H ]SCH 466036 was obtained at a
3
radiochemical purity of 98%, at specific activity of 82.9 Ci/mmole.
13
2
15
[
C, H , N]SCH 466036 was prepared in two steps as shown
3
in Figure 4.
The formation of the oxime 8 was accomplished by refluxing
15
ketone 7 with [ N]hydroxylamine hydrochloride in pyridine. This
led to a 50:50 mixture of the E and Z isomers, which were then
Results and discussion
3
Low specific activity [ H]SCH 466036, requested for pilot
absorption, distribution, metabolism and excretion studies, was
prepared by Raney nickel-catalysed exchange of ketone
5
intermediate 2 with 50 Ci/mL tritiated water (Figure 1).
Merck Research laboratories, Labeled Compound Synthesis, Department of
A batch of 29 mCi of crude compound 3 was isolated from the Process Chemistry, 126 E. Lincoln Avenue, RY 80R, Rahway, NJ 07065, USA
tritiation. After silica gel chromatography, the purified
*
Correspondence to: David Hesk, Merck Research Laboratories, Labeled
intermediate 3 was heated with methoxyamine hydrochloride in
pyridine to generate 39% of the desired E oxime 4. After
Compound Synthesis, Department of Process Chemistry, 126 E. Lincoln Avenue,
RY 80R, Rahway NJ 07065, USA.
reverse-phase high performance liquid chromatography (HPLC), E-mail: david.hesk@merck.com
J. Label Compd. Radiopharm 2015, 58 36–41
Copyright © 2015 John Wiley & Sons, Ltd.

D. Hesk et al.
3
Figure 1. Synthesis of [ H]SCH 466036.
1
3
3
Figure 2. H and H NMR of [ H]SCH 466036.
3
Figure 3. Synthesis of [Me- H
3
]SCH 466036.
13
2
methylated with [ C, H ]methyl iodide with potassium temperature melt, and the product is isolated by distillation.
3
hexamethyldisilylazide as the base. After silica gel As such, this would be a difficult reaction to replicate on
1
4
chromatography and chiral HPLC purification of the desired E the small scale that C synthesis is normally performed.
13
2
15
isomer, a batch of 114 mg of [ C, H , N]SCH 466036 was The latter reactions typically produce mixtures of the 2 and
3
isolated in overall 16% yield. Although a chiral column is not 4 isomers, although some control of the isomer ratio is
required for the separation of the oxime isomers, it provides a possible by careful control of the reaction conditions.
greatly improved separation over silica gel and a greatly Selective 2-cyanation has been reported by Vorbruggen and
1
2
improved loading over an achiral reverse-phase HPLC column.
Krolikiewicz, who used trimethylsilyl chloride and a 4.5
1
4
13,14
[
C]SCH 466036 was prepared as shown in Figure 5.
molar excess of sodium cyanide, and by Fife and Boyer,
2
-Cyanopyridines have classically been prepared by who reported the use of dimethylcarbamyl chloride with
8
reaction of copper cyanide with 2-bromopyridine and by potassium cyanide to selectively prepare 2-cyanopyridine
reaction of pyridine-N-oxide with dimethyl sulfate or by under mild conditions in high yields. Hence, pyridine-N-oxide
9
treatment of 1-methyloxypyridinium iodide with potassium was pretreated with dimethylcarbamyl chloride to form the 1-
1
0,11
cyanide.
The former reaction is conducted as a high dimethylaminocarbonyloxy pyridinium cation that was then
J. Label Compd. Radiopharm 2015, 58 36–41
Copyright © 2015 John Wiley & Sons, Ltd.
www.jlcr.org

D. Hesk et al.
1
3
2
15
3
Figure 4. Synthesis of [ C, H , N]SCH 466036.
1
4
treated with aqueous [ C]potassium cyanide at room hydroxybenzotriazole (HOBt) followed by removal of the
1
4
temperature to give compound 10. As [ C]2-cyanopyridine is butyloxycarbonyl group with trifluoroacetic acid and HPLC
1
4
volatile, the product was isolated by ether extraction and purification, a 13.5 mCi batch of [ C]SCH 466036 at a specific
thoroughly dried over anhydrous sodium sulfate prior to activity of 57.1 mCi/mmole was prepared in overall 13% yield
1
4
concentration to a volume of about 1 mL via distillation. The from [ C]potassium cyanide. The radiochemical purity as
ether solution containing compound 10 with 97 mCi (89% yield) determined by reverse-phase HPLC was >99%.
1
5
was then directly reacted with the Grignard reagent 11 to
generate 12 in 35% yield after purification. After treatment with
ethyl chloroformate to generate carboethoxy intermediate 13,
acidic hydrolysis yielded 14 as a di-hydrochloride salt in 76%
Experimental
yield. An attempt to directly convert 12 to 14 using 1- Materials
1
6
chloroethylchloroformate led to a 30% yield of 14. Formation
of the oxime E/Z mixture was accomplished by reaction with
methoxyamine hydrochloride under aqueous conditions with
careful adjustment of the pH to pH 3.5–5 with sodium
hydroxide. Treatment with 6 M hydrogen chloride in
isopropanol and heating of the resulting slurry of the
1
4
[
C]Potassium cyanide was purchased from Quotient Bioresearch,
3
Fordham Cambridgeshire, UK. [ H]Methyl iodide was purchased from
American Radiolabeled Chemicals Inc., St. Louis, MO, USA. [ C, H ]
3
Methyl iodide and [ N]hydroxylamine were purchased from Cambridge
Isotopes Inc., Tewksbury, MA, USA. Unlabelled intermediates 2, 5, 7 and
13
2
15
1
6 were obtained from Merck Process Chemistry (Union NJ, USA). All
dihydrochloride salt in isopropanol led to 90% conversion to reagents and solvents were purchased from Aldrich (Milwaukee, MI,
the desired E isomer 15. Finally, after a standard coupling with USA) and used as received unless otherwise stated. All synthetic steps
1
-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and were carried out under an atmosphere of argon.
1
4
Figure 5. Synthesis of [ C]SCH 466036.
www.jlcr.org
Copyright © 2015 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2015, 58 36–41

D. Hesk et al.
Liquid scintillation counting
capped with a rubber septum, frozen in liquid nitrogen, evacuated and
sealed in a flame. The reaction was heated for 24 h at 110 °C, before
being cooled to room temperature and opened. The ampoule contents
were partitioned between 1 M potassium hydroxide solution (2 mL) and
methylene chloride (5 mL). The methylene chloride layer was removed,
and the aqueous layer further extracted with methylene chloride
Radioactivity measurements were performed on a Packard 2900CA (Perkin
Elmer, Downers Grove IL, USA) liquid scintillation analyser using Scintiverse
BD (Perkin Elmer, Downers Grove IL, USA) as liquid scintillation cocktail.
(
3 × 5 mL). The combined extracts were washed with water (2 mL) and
Thin layer chromatography
evaporated to dryness. A total of 29 mCi of 3 at a radiochemical purity
of 79% (TLC system 2) was isolated. The crude product was purified by
chromatography on silica gel using a methylene chloride : methanol
gradient (0–10%). Fractions containing the purified product were pooled
and evaporated to dryness to yield 19 mCi of 3, which was used directly
in the next step.
Radio-thin layer chromatography (TLC) was performed with Whatman
LK6DF (silica gel 60) 5 × 20 cm, 0.25 mm plates. The plates were scanned
on a Bioscan 1000 linear analyser (Washington DC, USA).
System 1: Methylene chloride : methanol (90:10).
System 2: Methylene chloride:2M methanolic ammonia (90:10).
System 3: Ethyl acetate : hexane (50:50).
System 4: Methylene chloride:2M methanolic ammonia (95:5).
System 5: Methylene chloride:methanolic ammonia (98.5:1.5).
3
[
(
H](E)-(1-((2-Aminopyridin-4-yl)methyl)piperidin-4-yl)(4-
3
(methoxyimino)(pyridin-2-yl)methyl)piperidin-1-yl)methanone [ H]
SCH 466036 (4)
To
a solution of 19 mCi of 3 in pyridine (0.5 mL) was added
methoxyamine hydrochloride (8.8 mg). The reaction was heated at 90 °C
overnight and then assayed in HPLC system 2, which showed 80%
conversion to a (1:1) mixture of E and Z oxime isomers. The reaction
was evaporated to dryness, diluted with water (2 mL) and adjusted to
High performance liquid chromatography
A Waters 2695 Alliance System with a Waters 996 Photodiode array
detector (Milford MA, USA) was used. Radiochemical purity was
determined by a Radiomatic C150 radio flow detector (Perkin Elmer,
Downers Grove IL, USA) with Flo-Scint III liquid scintillation cocktail
>pH 10 with concentrated ammonia. The resulting suspension was
extracted with methylene chloride (3 × 5 mL). The combined extracts
were washed with water (2 mL), dried over anhydrous sodium sulfate
and evaporated to dryness. The product was purified by reverse-phase
HPLC using a 250 × 9.4 mm Extend C18 column with a mobile phase of
(Perkin Elmer, Downers Grove IL, USA). The following systems were used:
System 1: Agilent Zorbax Extend C18, 50 × 4.6 mm, 1.8 μ
66 nm, 0.05 M pH 10 triethylammonium acetate :
0
4
.05 M pH 9 triethylammonium acetate : acetonitrile (70:30) at a flow of
mL/min and 254 nm. Fractions containing the purified product were
2
acetonitrile (72:28) for 6 min followed by a step
gradient to acetonitrile, 1 mL/min.
pooled, evaporated to dryness and dissolved in 20 mL ethanol. A total
3
of 5.6 mCi of [ H]SCH 466036 4 was isolated at a radiochemical purity of
System 2: Agilent Zorbax Extend C18, 150 × 3 mm, 5 μ 254 nm,
9
9.8% and 99.0% (HPLC system 1 and TLC system 2) and a specific activity
3
0
.05 M pH 9 triethylammonium acetate : acetonitrile of 372 mCi/mmol. H NMR: (426 MHz) d -DMSO, δ8.56(d) 6%, δ7.78(d)
6
(
70:30) for 15 min followed by a step gradient to 51%, δ3.56(m) 43%.
acetonitrile, 0.5 mL/min.
System 3: YMC Pro C18 150×4.6mm, 5μ, 275 nm, 0.05 M pH 4
triethyl ammonium acetate (85:15) acetonitrile for
3
Synthesis of [Me- H ]SCH 466036
3
3
(
3
E)-(1-((2-Aminopyridin-4-yl)methyl)piperidin-4-yl)(4-(([Me- H ]
methoxyimino)(pyridin-2-yl)methyl)piperidin-1-yl)methanone (6)
1
1
5 min followed by a step gradient to acetonitrile,
mL/min.
System 4: Waters Xterra C18 150 × 3 mm, 5 μ, 230 nm, 0.02 M
Compound 5 (4.05 mg, 0.988 μmol) was suspended in potassium
sodium tetraborate : acetonitrile (85:15), 0.5 mL/min. hydroxide solution (0.1 M, 250 μL), water (250 μL) and methylene
chloride (300 μL). A stock solution of Aliquat 336 (670 mg) in toluene
(
10 mL) was prepared and 60 μL, 0.996 μmol of this was added to the
Specific activity measurements
3
reaction. A toluene solution of 85 Ci/mmol [ H]methyl iodide (100 mCi,
3
14
The chemical concentrations of [ H] and
[
C]SCH 466036 were 500 μL) was added, and the resulting two-phase mixture was vigorously
determined using an ultraviolet-based HPLC assay (systems 1 and 2)
stirred overnight. The reaction was diluted with the addition of water
using an authentic standard of SCH 466036 for [ H]SCH 466036 and a (2 mL) and ethyl acetate (2 mL). The ethyl acetate layer was removed,
3
3
14
reference standard of SCH 466036 for [Me- H
3
]SCH 466036 and [ C] and the aqueous layer extracted with ethyl acetate (3 × 3 mL). The
SCH 466036. These data were used in conjunction with the radiochemical
concentration to calculate the specific activity values.
combined ethyl acetate fractions were washed with water (1 mL), dried
over anhydrous sodium sulphate and evaporated to dryness to yield
6
4 mCi of crude product at 77% radiochemical purity in HPLC system 1.
The product was purified as described for 4 to yield 46 mCi at a
radiochemical purity of 99.0% and 98.4% (HPLC system 1 and TLC system
2) and a specific activity of 82.9 Ci/mmol.
Mass spectrometry
Mass spectra were acquired on the JEOL MStation magnetic sector mass
spectrometer (JEOL USA, Inc., Peabody, MA, USA) operating in the
positive-ion electrospray ionization or fast atom bombardment (FAB)
ionization mode.
13
2
15
Synthesis of [ C, H , N]SCH 466036
3
1
5
(
E)-(1-((2-Aminopyridin-4-yl)methyl)piperidin-4-yl)(4-((hydroxy[ N]
3
Synthesis of [ H]SCH 466036
imino)(pyridin-2-yl)methyl)piperidin-1-yl)methanone (8)
1
5
3
Compound 7 (1.0 g, 2.45 mmol) and [ N]hydroxylamine hydrochloride
1.04 g, 14.72 mmol) were dissolved in pyridine (12 mL). The reaction
was heated at 80 °C for 5 h at which point the reaction was complete
[
H](1-((2Aminopyridin-4-yl)methyl)piperidin-4-yl)(4-
(
picolinoylpiperidin-1-yl)methanone (3)
A 40 mg of Raney Ni slurry in acetone was weighed into a thick-walled (TLC system 1). The solution was evaporated to dryness, dissolved in
glass tritiation ampoule, and the acetone was removed under a stream toluene and re-evaporated to dryness. The crude product was initially
of nitrogen. Compound 2 (20 mg) was then added followed by dioxane purified by chromatography on silica gel using a gradient of methylene
(100 μL) and tritiated water (50 Ci/mL, 500 mCi, 10 μL). The tube was
chloride and methanol (0–50%). Fractions containing the purified
J. Label Compd. Radiopharm 2015, 58 36–41
Copyright © 2015 John Wiley & Sons, Ltd.
www.jlcr.org

D. Hesk et al.
14
product were pooled and evaporated to dryness to yield the purified Piperidin-4-yl(pyridin-2-yl)[ C]methanone dihydrochloride (14)
product 8, 1 g. The batch was further purified on a Chiralpak AD column,
A solution of 12 (39 mCi, 0.64 mmol) in anhydrous toluene (2 mL) was
50 cm × 5 cm, ID, with a mobile phase of hexane : isopropanol :
heated to 70 °C. Ethyl chloroformate (190 μL, 1.99 mmol) was added
dropwise, and the reaction heated for 2 h at this temperature. Analysis
by TLC system 5 showed complete reaction. The reaction was cooled to
room temperature, and potassium bicarbonate solution (1 M, 220 μL) added.
The organic layer was removed, and the residual aqueous layer extracted
with toluene (5 mL). The combined organic layers were dried over anhydrous
sodium sulfate, filtered and concentrated to yield 13 (29.5 mCi) as an oil.
HCl (6 M, 5.5 mL) was added, and the reaction heated overnight at 110 °C.
After cooling to room temperature, the reaction was concentrated
to a volume of around 1 mL and used directly in the next step.
diethylamine (75:25:0.5) at a flow of 40 mL/min and detection at
54 nm. Fractions containing the purified product were pooled and
evaporated to dryness to yield 440 mg (43%) of compound 8.
2
1
3
2
(
E)-(1-((2-Aminopyridin-4-yl)methyl)piperidin-4-yl)(4-(([ C, H ]
3
15
methoxy[ N]imino)(pyridin-2-yl)methyl)piperidin-1-yl)methanon
1
3
2
15
[
C, H , N]SCH 466036 (9)
3
Compound 8 (296 mg, 0.7 mmol) was dissolved in freshly distilled
dimethylformamide (DMF) (3.8 mL) and cooled to À10 °C. Potassium
14
hexamethyldisilazide solution (0.5 M, 1.4 mL, 0.7 mmol) was added (E)-Piperidin-4-yl(pyridin-2-yl)[ C]methanone O-methyl oxime (15)
dropwise over 5 min, and the solution was stirred at this temperature
13
2
The 1-mL solution of 14 generated in the previous step (29.5 mCi and
for 30 min. A stock solution of [ C, H
prepared in freshly distilled tetrahydrofuran (THF) (2 mL), and 0.46 mL
0.74 mmol) of this solution was added dropwise over 5 min to the
3
]methyl iodide (0.2 mL) was
0
2
.52 mmol) was diluted with water (2 mL) and basified to pH 10 with
5% sodium hydroxide solution. Methoxyamine hydrochloride solution
(
(
2
30 wt.%, 80 μL, 1.04 mmol) was added, and the pH was adjusted with
5% sodium hydroxide solution to a range of pH 3.5–5. The reaction
reaction. The reaction was stirred for 1 h at this temperature and then
quenched by the addition of brine (8 mL). The product was extracted
with methylene chloride (2 × 10 mL) and 10% ethanol in methylene
chloride (2 × 15 mL). The combined organic extracts were dried over
anhydrous sodium sulfate, filtered and evaporated to dryness. The crude
product was purified on silica gel and by HPLC on a ChiralPak AD column
using the same conditions described for compound 8 to yield a total of
was then heated at 55 °C for 5 h at which point monitoring by HPLC
system 4 showed complete conversion to a (1:1) mixture of E and Z
isomers of 15. The reaction was cooled to room temperature, basified with
2
5% sodium hydroxide solution to pH 13 and extracted with methylene
chloride (3 × 6 mL). The combined methylene chloride extracts were dried
over anhydrous sodium sulfate, filtered and evaporated to an oil. The oil
was then dissolved in toluene (280 μL), and a solution of HCl in
isopropanol (5–6 M, 280 μL) was added. The solution was heated to 65 °
C at which point a slurry was formed. After cooling to room temperature,
the solid was washed with isopropanol (3 × 1 mL) with the isopropanol
removed by a medium porosity filter stick. An additional 700 μL of
isopropanol was added, and the slurry was heated at 80 °C overnight.
Analysis by HPLC system 2 showed that a (90:8) ratio of E to Z oxime
had formed. The isopropanol was removed by filter stick, and the solid
dried under vacuum to yield 108 mg, 25 mCi of 15.
200 mg of compound 9. The batch was finally recrystallized from
methanol : water to yield a total of 114 mg (37% yield of 9). Purity (HPLC
system 2): 99.3%. FAB-MS/MS: m/z 442. High resolution MS: m/z 442.2864
13
2
15
3 2 5
confirms the composition of C23CH30H O N N.
1
4
Synthesis of [ C]SCH 466036
1
4
[
C]2-cyanopyridine (10)
In a flame-dried 10-mL flask, pyridine-N-oxide (180.3 mg, 1.896 mmol)
1
4
was added and heated to 70 °C until a complete melt was achieved. N, (E)-tert-Butyl (4-((4-(4-((methoxyimino)(pyridin-2-yl)[ C]methyl)
N-dimethylcarbamyl chloride (190 μL, 2.1 mmol) was added dropwise at piperidine-1-carbonyl)piperidin-1-yl)methyl)pyridin-2-yl)
this temperature, and a precipitate formed. Heating was continued for carbamate (17)
a further 2 h, and then the reaction was cooled to room temperature.
Compound 17 (125.7 mg, 0.37 mmol) and 15 (25 mCi, 108 mg and
The activated pyridine was dissolved in water (0.8 mL), and this solution
14
0.37 mmol) were dissolved in DMF (650 μL), and N-methyl morpholine
190 mL, 1.7 mmol), EDC (106.4 mg, 0.56 mmol) and HOBt (75 mg,
.56 mmol) were added. The reaction was stirred overnight at room
temperature and was shown to be complete by TLC system 1. The DMF
was removed by evaporation, and the resulting oil partitioned between
methylene chloride (10 mL) and water (4 mL). The methylene chloride
layer was removed, and the aqueous layer extracted with a further
was added dropwise to a 0 °C solution of [ C]potassium cyanide
123.5 mg, 100 mCi, 1.896 mmol). The reaction was stirred at this
(
0
(
temperature for 5 min and then at room temperature overnight. Analysis
by TLC system 3 showed complete reaction. Potassium carbonate
solution (1 M, 200 μL) was added to the reaction, and the product was
extracted with ether (3 × 7 mL). The combined ether extracts were
washed with water (1 mL), dried over anhydrous sodium sulfate, filtered
and concentrated by distillation using a short path condenser to a
volume of about 1 mL and used directly in the next step. Yield = 97 mCi.
1
0 mL of methylene chloride. The combined methylene chloride layers
were washed with sodium bicarbonate solution (0.3 M, 5 mL), dried over
anhydrous sodium sulfate, filtered and evaporated to yield 17 (18 mCi) as
an oil, which was used directly in the next step.
1
4
(
1-Methylpiperidin-4-yl)(pyridin-2-yl)[ C]methanone (12)
14
(
(
[
E)-(1-((2-Aminopyridin-4-yl)[ C]methyl)piperidin-4-yl)(4-
(methoxyimino)(pyridin-2-yl)methyl)piperidin-1-yl)methanone
14
The ether solution of [ C]2-cyanopyridine (97 mCi) was cooled to 0 °C,
1
5
and a THF solution of (1-methylpiperidin-4-yl)magnesium chloride 11
2.5 M, 1.5 mL, 3.84 mmol) was added dropwise. The reaction was stirred
14
C]SCH 466036 (18)
(
at 0 °C for 90 min and then was quenched by the addition of
concentrated hydrochloric acid (HCl) (420 μL), crushed ice and ethyl
acetate (4 mL). Stirring was continued for a further 20 min at 0 °C and
at room temperature for 2 h at which point analysis by TLC system 4
showed complete reaction. The ether layer was removed, and the
aqueous fraction was basified with concentrated ammonia to pH 11
and extracted with ethyl acetate (3 × 10 mL). The combined organic
extracts were washed with brine (2 mL), dried over anhydrous sodium
Trifluoroacetic acid (500 μL) was added to a solution of 17 (18 mCi,
0.327 mmol) in methylene chloride (1.8 mL), and the reaction stirred
overnight at room temperature. Analysis by TLC system 1 showed
complete reaction. Water (2 mL) was added, and the reaction basified
to pH 10 with 25% sodium hydroxide solution. The methylene chloride
layer was removed, and the aqueous layer extracted with methylene
chloride (3 × 7 mL). The combined methylene chloride layers were
washed with water, dried over anhydrous sodium sulfate, filtered and
sulfate, filtered and evaporated to an oil. The crude product was purified evaporated to an oil. The crude product was purified by chromatography
by chromatography on silica gel using a gradient of methylene chloride :
methanol (0–5% methanol) to yield 12 (39 mCi) as a yellow oil after
concentration, which was used directly in the next step.
on silica gel using a methylene chloride : 2 M methanolic ammonia
gradient (0–5% methanolic ammonia). Fractions containing the purified
product were pooled and evaporated to dryness to yield 16 mCi of 18,
www.jlcr.org
Copyright © 2015 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2015, 58 36–41

D. Hesk et al.
which contained 5% of the undesired Z oxime isomer. The batch was
further purified by reverse-phase HPLC as described for 3 to yield a total
of 13.5 mCi at a radiochemical purity of 99.1% and 99.6% (HPLC systems
[2] W. Wu, H. Liao, D. J. S. Tsai, PCT Int. Appl. 2003, 27pp, WO
2003033488.
[
3] R. G. Aslanian, W. G. Tom, X. Zhu, PCT Int. Appl. 2006 44pp, WO
006078775.
4] R. G. Aslanian, J. E. Lachowicz, M. Y. Berlin, J. J. Hwa, PCT Int. Appl.
008, 179pp, WO 2008108957.
[5] D. Hesk, C. F. Lavey, P. McNamara, J. Labelled Comp. Radiopharm.
010, 53, 722–730.
2
2
and 3) and a specific activity of 57.1 mCi/mmol.
[
2
Acknowledgements
2
[
6] J. Garnett, M. Long,Catalytic exchange methods of hydrogen
isotope labelling. In Isotopes in the Physical and Biomedical
Sciences (Eds.: 1, J. R. Jones, E. Buncel), Elsevier: Amsterdam,
The authors wish to thank Dr Ingrid Mergelsberg from Merck
Process Chemistry for providing intermediates and for helpful
discussions. Thanks are also due to Dr T. M. Chan and Ms Rebecca
Osterman for the nuclear magnetic resonance analysis and to Dr
Pradip Das for the mass spectrometry analysis of the final compounds.
1
987, pp. 86–121.
3
[
7] E. A. Evans, D. C. Warrel, J. A. Elvidge, J. R. Jones, Handbook of H NMR
Spectroscopy and Applications, Wiley, Chichester, 1985.
[
8] L. C. Craig, J. Am. Chem. Soc. 1934, 56, 231–232.
[
9] W. E. Feeley, E. M. Beavers, J. Am. Chem. Soc. 1959, 81, 4004–4007.
Conflict of Interest
[
[
10] T. Okamoto, H. Tani, Chem. Pharm. Bull. 1959, 7, 130–131.
11] W. D. Crow, A. N. Khan, M. N. Paddon-Row, D. S. Sutherland, Aust. J.
Chem. 1975, 28, 1763–1773.
The authors did not report any conflict of interest.
[
[
[
12] H. Vorbruggen, K. Krolikiewicz, Synthesis 1983, 4, 316–319.
13] W. K. Fife, J. Org. Chem. 1983, 48, 1375–1377.
14] W. K. Fife, B. D. Boyer, Heterocycles 1984, 22, 1121–1124.
References
[
1] R. G. Aslanian, N. Y. Shih, P. Ting, M. Y. Berlin, S. Rosenblum, K.
McCormick, W. C. Tom, C. W. Boyce, P. Mangiaracina, M. W. Mutahi,
J. J. Piwinski, PCT Int. Appl. 2002, 144pp, WO 2002032893.
[15] Prepared from 4-chloro-1-methylpiperidine, Mg, THF.
[16] R. A. Olofson, J. T. Martz, J. P. Senet, M. Piteau, T. Malfroot, J. Org.
Chem. 1984, 49, 2081–2082.
J. Label Compd. Radiopharm 2015, 58 36–41
Copyright © 2015 John Wiley & Sons, Ltd.
www.jlcr.org